Single-molecule imaging reveals the mechanism of Exo1 regulation by single-stranded DNA binding proteins
Logan R. MylerIluminada GallardoYi ZhouFade GongSoo-Hyun YangMarc S. WoldKyle M. MillerTanya T. PaullIlya J. Finkelstein
94
Citation
81
Reference
10
Related Paper
Citation Trend
Abstract:
Significance Exonuclease 1 (Exo1) is a conserved eukaryotic nuclease that participates in DNA repair and telomere maintenance. Here we use high-throughput single-molecule imaging to examine Exo1 activity on DNA and in the presence of single-stranded DNA binding proteins. We report that both human and yeast Exo1 are processive nucleases but are rapidly turned over by replication protein A (RPA). In the presence of RPA, Exo1 retains limited DNA-processing activity, albeit via a distributive binding mechanism. This rapid turnover by RPA can appear stimulatory or inhibitory in gel-based assays, clarifying conflicting results in the existing literature. RPA-depleted human cells show elevated Exo1 loading but reduced overall DNA resection, underscoring the many roles of RPA in regulating DNA resection in vivo.Keywords:
Replication protein A
Nuclease
DNA clamp
Rad50